Applications

for AbFlex BMI-1 antibody (rAb)Application Notes

AbFlex™ recombinant antibodies are genetically derived from DNA sequences of parental hybridoma clones. For details on the parental clone, see Catalog No. 39993.

Note: many chromatin-bound proteins are not soluble in a low salt nuclear extract and fractionate to the pellet. Therefore, we recommend a High Salt / Sonication Protocol when preparing nuclear extracts for Western Blot.

for AbFlex BMI-1 antibody (rAb)Buffer

for AbFlex BMI-1 antibody (rAb)References

Western blot of AbFlex® BMI-1 antibody (rAb).20 µg of K562 cell nuclear extract was run on SDS-PAGE and probed with AbFlex® BMI-1 antibody.

AbFlex® BMI-1 recombinant antibody (rAb) tested by ChIP-SeqChromatin immunoprecipitation (ChIP) was performed using the ChIP-IT® High Sensitivity Kit (Cat. No. 53040) with 30 µg of mouse embryonic fibroblast (MEF) chromatin and 4 µg of BMI-1 antibody. ChIP DNA was sequenced on the Illumina NextSeq and 9.9 million sequence tags were mapped to identify BMI-1 binding sites on chromosome 2.

for AbFlex BMI-1 antibody (rAb)Background

AbFlex® antibodies are recombinant antibodies (rAbs) that have been generated using defined DNA sequences to produce highly specific, reproducible antibodies. Each AbFlex antibody contains a 6xHis-Tag, an avidin tag sequence for enzymatic biotin conjugation using the biotin ligase, BirA, and a sortase recognition motif (LPXTG) to attach a variety of labels directly to the antibody including fluorophores, enzymatic substrates (HRP, AP), peptides, drugs as well as solid supports.
BMI-1 is a member of the Polycomb PRC1 complex that is recruited to transcriptionally repressed genes subsequent to histone H3 lysine 27 methylation in order to maintain repression. In the PRC1 complex, BMI-1 is required to stimulate the E3 ubiquitin-protein ligase activity of RNF2/RING2, resulting in the ubiquitylation of histone H2A at lysine 119. BMI-1 acts as an oncogene and cooperates with c-myc in the initiation of lymphoma. High levels of expression of BMI-1 are observed in metastatic melanoma, and BMI-1 has been implicated in several other cancers. BMI-1 is required for hematopoietic stem cell self renewal.

Chromatin IP Assays

This antibody has been validated for use in ChIP and/or ChIP-Seq, and can be used with Active Motif's ChIP-IT® High Sensitivity Kit or our magnetic bead-based ChIP-IT® Express Kits. For an overview of all of our ChIP products, please visit our Chromatin IP Page to learn about ChIP products designed for use with AbFlex® BMI-1 antibody (rAb).

Bridging Antibody to Improve ChIP, MeDIP and IP

Some isotypes of mouse IgG antibodies do not bind well to protein G-conjugated agarose/magnetic beads, which are commonly used in ChIP, MeDIP and IP experiments. Capture efficiency can be improved by including an anti-mouse IgG bridging antibody because it binds with a strong affinity to both the protein G beads and the mouse IgG antibody. By increasing the affinity of the mouse primary antibody for the protein G bead, the Bridging Antibody for Mouse IgG can help improve the results of all ChIP, MeDIP and IP experiments that use AbFlex® BMI-1 antibody (rAb).

for AbFlex BMI-1 antibody (rAb)Storage

Some products may be shipped at room temperature. This will not affect their stability or performance. Upon receipt, unconjugated antibodies may be stored at -20°C for up to 2 years. Fluorophore- & enzyme-conjugated antibodies should be stored at 4°C. Fluorophore-conjugated antibodies should be protected from light. Keep reagents on ice when not in storage; to avoid repeated freeze/thaw cycles, we recommend aliquoting items that will be stored frozen into single-use fractions prior to freezing.

for AbFlex BMI-1 antibody (rAb)Guarantee

This product is guaranteed for 6 months from date of receipt.

This product is for research use only and is not for use in diagnostic procedures.